The invention relates to a method and device for determining the presence of an antigen in test samples through the use of a specially designed enzyme labeled "double site" immunoassay technique.
Enzyme labeled immunoassay employing antibodies which react with different domains on the same antigen have been described in the scientific literature. In general, all such "double site" immunoassays are conducted as follows: One member of the antibody pair is fixed to a solid support such as the wall of a plastic tube or the surface of a plastic stick. The second antibody is conjugated with an enzyme label. The antigen containing sample and the second antibody conjugate are mixed together with the first antibody bound to the solid support. In the presence of the antigen the antibody conjugated becomes bound to the solid support via the opposite member of the antibody pair.
The solid support is then washed thoroughly to remove any unbound conjugate. Finally the solid support is incubated with a color forming enzyme substrate. The color reaction can then be read visually or with an instrument to indicate the presence and quantity of the antigen. Such two site immunoassays can be specific and rapid but require multiple washing and color forming incubation steps.
The present invention is a two site immunoassay which has no washing or separate color reaction steps. The antigen containing sample is mixed with the two antibody reagents and the mixture is applied to the surface of a two layered test device. The bottom layer contains the color forming substrate. In the presence of the antigen the conjugate is carried to the bottom layer and produces a color reaction. In the absence of the antigen all the conjugate is trapped in the solid phase top trapping layer and fails to reach the lower color forming layer. The principle of the invention is exactly the opposite of the previous two site immunoassays, because the conjugate which fails to bind the antigen is trapped on the solid phase. The conjugate which binds to the antigen washes through the solid phase zone to the color forming layer.
The foregoing and other advantages of the instant invention will become apparent to those skilled in the art from a reading of the following specification and claims.